A program of research is proposed to elucidate the control mechanisms for folate metabolism in the developing and adult rat liver, and the changes which occur in regenerating liver, and in a series of hepatomas of different growth rates. Attempts will be made to establish the relationship between the multiple forms of dihydrofolate reductase and to determine the pattern across the hepatoma spectrum, using polyacrylamide gel electrophoresis. We will also investigate the way in which the balance is maintained between the different routes of utilization for various tetrahydrofolate cofactors; this will require a study of total activity, chemical and kinetic properties, and inhibition profiles of at least seven folate enzymes. Folate cofactor pool levels, one-carbon group donor concentrations, NADP ion and NADPH concentrations will be measured in freeze-clamped liver and hepatoma preparations. We will study the possibility that susceptibility of folate enzymes to physiological control may be lost or modified in hepatomas. The principle sources of one-carbon groups in liver and hepatomas will be established, and the way in which one-carbon groups from various sources are distributed between the different acceptor compounds will be studied by isotope distribution methods. A program of experimental chemotherapy will be designed to exploit changes which are found in hepatoma folate biochemistry. The effects of anti-metabolites will be studied on isolated, reconstituted multienzyme systems, on hepatomas growing in tissue culture, and as solid tumors in the whole animal. Mathematical models will be used to predict optimal conditions for tumor response to drug treatment; by using the knowledge of hepatoma folate biochemistry obtained in this study, drug toxicity to hepatomas may be maximized while toxicity to liver and host proliferative tissues will be minimized. Drug combinations (or drug plus metabolic intermediary combinations) predicted on biochemical grounds to have the highest therapeutic index will be subjected to detailed pharmacological evaluation.